Impact of Human Papillomavirus Vaccination on Male Disease: A Systematic Review

Human papillomavirus (HPV)-related diseases are highly prevalent in men worldwide, comprising external anogenital condyloma, anal intraepithelial neoplasia (AIN), penile intraepithelial neoplasia (PIN), and anogenital and oropharyngeal cancers. There is exceptionally low vaccine coverage in the male population. Only 4% of men were fully vaccinated, worldwide, as of 2019. The aim of this review is to assess the impact of HPV vaccination on male disease. Three databases (MEDLINE, Web of Science, Scopus) and Clinical Trials.gov were searched. We included thirteen studies, eight randomized controlled trials (RCTs), and five cohorts, comprising a total of 14,239 participants. Regarding anal disease, seven studies reported HPV vaccine efficacy ranging from 91.1% to 93.1% against AIN1, and ranging from 89.6% to 91.7% against AIN2|3 and anal cancer. Five studies showed an efficacy against genital condyloma of 89.9% in HPV-naïve males, varying between 66.7% and 67.2% in intention-to-treat populations. Studies reporting no efficacy have included older participants. These results support vaccination of young men previously infected, beyond HPV-naïve males. The evidence quality was moderate to low for most outcomes, namely genital diseases. RCTs are needed to assess the efficacy of HPV vaccination on male oropharyngeal cancer.


Introduction
Human papillomavirus (HPV) is the most common agent of sexually transmitted diseases [1]. HPV-related diseases have a high prevalence in men of all ages around the world [2]. Overall, 45 genotypes can specifically invade the anogenital tract and epithelium of the oral cavity, oropharynx, and larynx [3]. The most common genotypes in the male population are HPV-6, 11, 16, and 18 [2]. Benign lesions such as external anogenital warts and recurrent respiratory papillomatosis are caused mostly by low-risk types such as HPV-6 and HPV-11 [4,5]. Malignant anogenital and head and neck diseases are more associated with high-risk (HR)-HPV types, of which HPV-16 and HPV-18 are the most common [2]. HPV infection can resolve naturally without causing disease [6], and even in persistent infection, a large number of infected men do not present clinical manifestations [7]. However, when present, they may range from benign lesions such as anogenital warts or respiratory papillomatosis to more severe situations such as anal and genital cancers and their precursors, and head and neck cancer (HNC) [8]. Since the HPV vaccination of girls and women has shown a protective effect against HPV-related diseases [9], extending the vaccination to boys and men could lead to a lower burden of diseases.
The control group had to be males who received placebo or were unvaccinated. For a representative sample of the male population worldwide, we did not exclude men based on sexual orientation and on health status such as HIV status, HPV infection status, or history of HPV-related disease. We excluded (1) studies that evaluated immunogenicity focused on clinical manifestations of disease; (2) studies about infection incidence rates and prevalence, considering that we would not be able to analyze the presence or absence of a relationship between HPV vaccines and HPV-related diseases; (3) studies reporting administration of the vaccine combined with biologically active adjuvants; (4) the use of intralesional HPV vaccine; (5) studies portraying the efficacy of the vaccine by herd protection given by female HPV vaccination; and (6) estimates using statistical models to avoid the influence of confounders on the measurement of the direct efficacy of the vaccination; (7) reviews; (8) systematic reviews; (9) non-randomized controlled trials; (10) cross-sectional studies; (11) case series and (12) case reports; and (13) animal studies (Supplementary Material File S1).
Our search was focused on high-level scientific evidence of the efficacy of HPV vaccination on clinical manifestations of male disease. We considered studies published from January 2010 to October 2022. No study was excluded based on language.
The searched electronic databases were MEDLINE (PubMed), Web of Science, and Scopus, and were last accessed on 29 October 2022, for all studies. Additionally, ClinicalTrials. gov was searched systematically (last update 22 December 2022) for unpublished or ongoing trials. The queries used are presented in Supplementary Material File S1, and the terms "HPV", "human papilloma virus", "vaccination", "vaccine", "immunization", "efficacy", "effectiveness", and "male disease" were the search terms used for the review. Details on the search strategy are reported in Supplementary Material File S1.

Selection of Studies and Data Extraction
Screening and a full-text review of the searched studies were performed by two independent investigators (C.R. and C.L.) using the COVIDence platform (COVIDence Systematic Review software, Veritas Health Innovation, Melbourne, Australia), and conflicts were solved by reaching a consensus.
Two independent reviewers (C.R. and Â.R.F.) used standardized Excel tables to allow the extraction of relevant study data. Conflicts that emerged during the extraction were solved by a third reviewer (C.L.).
The variables considered important to data extraction included first author and publication year, study design, base study, study site, study period, duration of follow-up, characteristics at enrollment, intervention and comparator, number of participants, inclusion criteria, exclusion criteria, outcomes, results, limitations, sponsorships, and conflicts of interest.

Study Quality Assessment (Risk-of-Bias)
The risk-of-bias assessment was carried out using the following recommended tools: Cochrane risk-of-bias tool for randomized trials (RoB2) [28], and the risk-of-bias in nonrandomized studies for interventions (ROBINS-I) [29] for cohort-type studies. Two independent reviewers (Â.R.F. and C.L.) resolved any conflicts over risk-of-bias judgement by reaching a consensus. The RoB2 tool assesses five domains of bias in randomized controlled trials: (1) bias arising from the randomization process, (2) bias due to deviations from intended interventions, (3) bias due to missing outcome data, (4) bias in measurement of the outcome, and (5) bias in selection of the reported results. The overall risk-of-bias judgement was categorized either as low, some concerns, or high.
The ROBINS-I tool comprises seven domains of bias in non-randomized controlled studies: confounding factors, selection of participants for the study, classification of interventions, deviations from intended intervention, missing data, measurement of outcomes, and selection of the reported results. The overall ROBINS-I judgement was categorized as low, moderate, serious, or critical.
For both tools, the categorization of a domain in a level of risk-of-bias will classify the study at this severity score.

Glossary of Nomenclature
To homogenize the nomenclatures of the diseases and simplify the reading, we will refer to high-grade anal intraepithelial neoplasia (HGAIN) and high squamous intraepithelial lesion (HSIL) as anal intraepithelial neoplasia (AIN) grade 2/3, and low squamous intraepithelial lesion (LSIL) as AIN grade 1. Similarly, we will refer to high-grade penile intraepithelial neoplasia (HGPIN) and high squamous intraepithelial lesion (HSIL) as penile intraepithelial neoplasia (PIN) grade 2/3, and low squamous intraepithelial lesion (LSIL) as PIN grade 1 [30] (Supplementary Material File S2).

Studies' General Characteristics
Twelve studies used the 4vHPV vaccine and a single one used the 9vHPV vaccine [39]. All the included studies assessed the efficacy of the vaccine against disease caused by vaccine types, except the study by Goldstone et al. [28], which failed to demonstrate the efficacy of 4vHPV vaccine against anal disease and EGL caused by ten additional non-vaccine HPV types.
Two studies evaluated the efficacy of the 4vHPV vaccine on HPV-naïve men [32,35]; one of them enrolled only men that were HPV-naïve to all four genotypes covered by the vaccine [32], while the other had the inclusion criterion of being seronegative for at least two vaccine genotypes [35]. Concerning HPV-related disease history, six studies evaluated vaccine efficacy in men with previous HPV-related diseases [35,36,38], and seven studies included men without history of disease. Abbreviations: 4vHPV vaccine = Quadrivalent HPV vaccine; AIN = Anal intraepithelial neoplasia; ASCC = Anal squamous cell carcinoma; CVG = Catch-up vaccination group; EVG = Early vaccination group; HGAIN = High-grade anal intraepithelial neoplasia; HIV = Human immunodeficiency virus; LGAIN = Low-grade anal intraepithelial neoplasia; LSIL = Low-grade intraepithelial lesion; MSM = Men who have sex with men; PCR = Polymerase chain reaction; RCT = Randomized controlled trial; STI = Sexually transmitted infection. ** Note: HGAIN and HSIL corresponds to AIN grade 2/3; LSIL corresponds to AIN grade 1; Penile HGAIN and HSIL corresponds to PIN grade 2/3; penile LSIL corresponds to PIN grade 1 [30]. ∆ Anal HPV-related diseases; # Genital HPV-related diseases.
We did not find studies that had assessed other disease outcomes, such as RRP or HNC, related to HPV in the male population.

Outcomes Reported in the Included Studies of HPV-Related Anal Disease
Ten of the included studies focus on anal male disease; data from these studies are summarized in Table 2.
Seven studies presented efficacy against anal HPV-related disease [17,31,32,[39][40][41][42]. Participants' age ranged from 16 to 26 years old, except in one study that had enrolled men older than 26 years [42]. Efficacy against anal condyloma acuminata was assessed by a cohort [42] that showed a significantly lower incidence rate in vaccinated participants, with an HR = 0.45. Two RCTs [31,32] and one cohort [41] assessed efficacy against AIN1. In the RCTs, efficacy ranged from 91.1% (including condyloma acuminata) [31] to 93.1% (not including condyloma acuminata) [32]. In the cohort study, a sub-study based on a previous RCT, the early and the catch-up vaccination groups (EVG and CVG, respectively) both had a lower incidence rate of anal disease when compared to the placebo group of the base study [41]. Efficacy against AIN2|3 was assessed using two RCTs; one RCT reported an efficacy of 91.7% against AIN2|3 [31], and the other one an efficacy of 89.6% or worse against AIN2|3 in the HPV-naïve population, and 50.3% in the intention-to-treat (ITT) population [32]. A cohort reported that vaccinated participants revealed a lower incidence rate and lower risk of recurrence of AIN2|3 at 1 year (HR = 0.42), and similar risks at years 2 and 3 (HR = 0.50, and HR = 0.52, respectively) [17]. Regarding anal cancer outcomes, only one cohort [41] reported a protective effect in vaccinated men, both in EVG and CVG groups. Of the aforementioned studies, four studies included male participants without history of HPV disease, and all of them were HIV-negative at enrollment [31,32,39,40]; two of them assessed efficacy on adolescents [39,40], and one of them included only MSM participants [31]. The other studied both heterosexual men and MSM [32]. Three studies enrolled men with history of disease, all HIV-negative men; two studies included only MSM [17,42], while a third one included both MSM and heterosexual men [41].
Three studies showed no efficacy; participants in these studies were ≥26 years old [33,35,36]. Three RCTs [33,35,36] assessed efficacy for AIN2|3, and none of them showed significant differences between vaccinated and unvaccinated male participants. Considering anal cancer outcomes, as assessed by two RCTs, none found significant differences between vaccinated men and control groups. Two studies [35,36] included MSM living with HIV infection (PLWHMSM) with history of HPV disease.  For CVG, the incidence per 10,000 person years was lower during the long-term follow-up period than during the base study period for AIN and anal cancer related to HPV-6/11/16/18. In addition, the incidence of anal disease in both the EVG and CVG during the long-term follow-up period was significantly lower than in placebo recipients during the base study

Outcomes Reported in the Included Studies of HPV-Related Genital Disease
Eight studies analyzed male genital diseases attributable to HPV; data are summarized in Table 3. We found five studies that reported efficacy [32,37,[39][40][41]; at enrollment, participants' ages ranged from 16 to 27 years old. Efficacy against genital condyloma acuminata and PIN1|2|3 was assessed by five studies, two RCTs [32,37], and three cohorts [39][40][41]. One RCT [33] reported an efficacy of 89.9% against genital condyloma for the HPV-naïve population, and 66.7% for the ITT population, while the other RCT [37] reported an efficacy of 67.2% in the ITT population. Concerning the PIN outcome, both RCTs found no significant differences between vaccinated and placebo groups [32,37]. All three cohort studies evaluated the same outcomes, genital condyloma and PIN, and reported a decrease in the incidence rate in vaccinated participants [39][40][41]. Regarding penile cancer, four studies assessed this outcome: an RCT [37] and three cohorts studies [39][40][41]. However, efficacy could not be assessed, as there were no cases diagnosed. The cohort studies reported low incidence rates. Four of the studies mentioned above [32,37,39,40] included HIV-negative men without a history of HPV disease; two studies within these enrolled adolescents [40,41]. Heterosexual men and MSM were eligible in two studies [32,41].
Three RCTs showed no efficacy of the HPV vaccine against genital condyloma [34,35,38]. Participants were 26 years or older at enrollment [35,38], except in one study [34] (age ranged from 16 to 26 years). One of the RCTs assessed PIN1 including condyloma and reported not having enough statistical power to evaluate both outcomes [34]. None of these studies assessed PIN2|3 or penile cancer. Two of the studies had participants with a history of HPV disease [35,38]; one included PLWH and MSM participants [35], while the other enrolled only HIV-negative heterosexual men [38]. One study included both heterosexuals and MSM participants without a history of HPV disease and with unknown HIV status [34].

Study Quality Assessment (Risk-of-Bias)
For RCT studies, topics concerning randomization after entry, blindness of treatment, and loss to follow-up are of major importance for risk-of-bias assessment. In cohort studies in which some factors are not controlled, evaluation of potential confounders, such as biases in participants' selection and measurement of outcomes, is of paramount significance.
Regarding risk of bias in this systematic review, the studies were categorized as having low, moderate, or high risk of bias (Tables 2 and 3). Five studies on anal HPV-related diseases were classified as having moderate risk of bias, and the remaining five as having low risk of bias. Concerning genital HPV-related diseases, three were categorized as having moderate risk of bias, and five as having low risk of bias.
Risk of bias is associated with different methodologies used between patients within groups, their blinding to treatment, suspension of the studies, the reliability of the diagnostic techniques applied, and the different times from entry to follow-up. Table 3. Overview of genital HPV-related outcomes reported in the studies.

Study
Outcomes Results

Limitations Sponsorship/Conflicts of Interest Risk-of-Bias
[37] Giuliano, A. R., 2011 Reduction in the incidence (as compared with placebo) of EGL associated with HPV present in 4vHPV vaccine or to any HPV type.
The ITT efficacy to reduce recurrent EGL was 60.2% overall; in particular, 4vHPV vaccine has 67.2% of efficacy in preventing condyloma. The PP analysis showed an overall 83.8% efficacy and 89.4% 4vHPV vaccine efficacy for condyloma. Penile cancer was not evaluated due to lack of cases in the sample, and a low number of participants presented PIN For the EVG, when compared to placebo group of the base study, results of incidence were null for external genital warts related to HPV-6/11 and external genital lesions related to HPV-6/11/16/18. For CVG, the incidence per 10,000 person years was lower during the long-term follow-up period than during the base study period for external genital warts related to HPV-6/11, and external genital lesions related to HPV-6/11/16/18. Comparing EVG and CVG in the present study, the incidence of the external genital warts related to HPV-6/11 and external genital lesions related to HPV-6/11/16/18 was similar to the incidence in the EVG

Discussion
We conducted a systematic review that assessed the efficacy of HPV vaccination in male disease. Overall, we found that HPV vaccine in males was effective, although the characteristics of the study participants and the outcomes assessed influenced the impact of vaccination upon HPV-related anogenital disease.
The participants' age and history of HPV-related disease influenced vaccination efficacy. We observed higher efficacy in men who were naïve to the respective HPV types evaluated in the studies. Interestingly, the seroprevalence of any HPV vaccine type increased with age [43,44]. Thus, the likelihood of being HPV-naïve is higher in childhood/early adolescence before exposure to HPV through sexual activity. Accordingly, the studies that assessed vaccine efficacy in males older than 26 years and with previous history of HPVrelated disease found no efficacy against genital disease [35,38]. The study by Goldstone S. et al. [41] reported a similar incidence of EGL in the EVG (young men aged 16 to 26) and in the CVG (men older than 26 years). However, the participants of the CVG were young men belonging to a placebo group in a previous RCT [37], and history of HPV-related genital lesions was one of the exclusion criteria. On the other hand, for anal disease, two studies assessed men with previous history of disease and showed efficacy [17,42], while two others reported no efficacy in these participants [35,36]. In fact, studies that aimed to assess the efficacy of 4vHPV vaccine in the prevention of recurrent AIN2|3 [17,36] reported different results. Swedish et al. [17] found a decreased risk of AIN2|3 recurrence in vaccinated individuals, while Gosens et al. [36] found no efficacy of the 4vHPV vaccine in the prevention of AIN2|3 recurrence. Notably, this last study enrolled HIV-positive men and the follow-up was very short (1.5 years) which might affect these findings. Wilkin et al. [33] evaluated the efficacy of the 4vHPV vaccine against anal HSIL in HIV-positive men aged 27 years or older; about a third of them had a history of anal disease, and no efficacy of 4vHPV vaccine was found. Therefore, these studies do not support the ability of the HPV vaccine in HIV-positive men aged 27 years or older to prevent or improve anal disease outcomes. HPV vaccination provides protection against persistent HPV infections and against the development of AIN2|3 in individuals without prior vaccine-type HPV infection [45]. However, catch-up vaccination is also recommended for MSM and HIV-infected individuals up to 26 years of age [46].
Regarding data in males aged between 16 and 26 years, (CVG) Ferris et al. [40] and Goldstone et al. [41] reported HPV vaccine efficacy against genital condyloma, PIN and AIN2|3, although the results were inferior to those of the younger group (EVG). Both studies were of high quality (and low risk of bias), thereby allowing generalizations to other populations and supporting the current recommendation for HPV vaccination of men up to age 26 years [47].
Regarding vaccine efficacy in the prevention of HPV-related anogenital diseases, the included studies found that the highest vaccine efficacy was observed for AIN 1|2|3 [17,31,32,[40][41][42], followed by genital condyloma [32,37,[39][40][41] and anal condyloma [31,[40][41][42]. The studies that assessed these outcomes have from high to fair quality (i.e., low to moderate risk of bias). In contrast, a single study [40] assessed vaccine efficacy against PIN. The authors did not find any protective effect of the HPV vaccine against HPV-6, 11, 16 and 18 related to PIN. However, this study has critical limitations: the small size of the study population, and collection of genital specimens in several centers without standardization of the procedure. In fact, male genital sampling has not been standardized as it has in women [48]. It should be noted that the retrieved studies couldn't assess vaccine efficacy/effectiveness against penile or anal cancer [37,[39][40][41] due to absence of cases present during the studies. The follow-up ranged from 2.9 years [37] to 10 years [39].
PLWH and MSM have a greater risk of developing disease associated with HPV. HIV infection can reduce vaccine immunogenicity and effectiveness due to low immune response in these patients [1]. MSM may be at greater risk of disease, as they do not benefit from the herd immunity provided by female vaccination against HPV.
Vaccinating women can protect heterosexual men, but this herd protection does not extend to MSM and bisexual men [49]. Not considering this group for vaccination can contribute to an increase HPV-related diseases, especially anogenital cancer [50,51]. Barriers to vaccine uptake in these groups do arise from costs, and from incorrect association of HPV vaccine with cervical cancer alone [52]. Additionally, a lack of physicians' recommendations, at times due to non-disclosure of patients' sexual orientation [53], contributes to low HPV vaccine coverage.
Our systematic review included studies that enrolled PLWH, although none of them reported on the difference in HPV vaccine efficacy between PLWH and HIV-negative people. The study by Bergman et al. and two systematic reviews by Zhan et al. found that HPV vaccination is effective and safe in PLWH [54,55]. However, they highlighted that vaccine efficacy in the prevention of HPV-related neoplasia remains unknown. In our review, the studies that showed efficacy against anal and genital HPV-related disease enrolled HIVnegative men. Three studies [35,37,38] included PLWH, and reported that vaccinated and unvaccinated men had the same risk of AIN2|3. Additionally, Gosens et al. [36] reported similar cumulative AIN2|3 recurrence between vaccinated men and the placebo group.
Our review supports previous findings, namely regarding higher HPV vaccine efficacy in boys before sexual debut [56]. Moreover, HPV vaccination in males is effective against AIN1|2|3 [57] and anogenital warts [57,58]. As shown in our systematic review, the body of evidence is much weaker regarding penile precancerous lesions in males.
One of the strengths of this systematic review results from the number of participants analyzed, which reached 14,239 boys and men. Another strength is the fact that we were able to include a diverse study population (MSM as well as heterosexual men, HIV-negative men and PLWH, HPV-naïve males and men with history of HPV-related disease) from several countries worldwide, meaning we could observe real-life scenarios and allowing the generalization of the findings. Our review was conducted based on internationally accepted methods, and the tools used for evaluation of the studies' risk-of-bias are part of the recommended ones. Concerning the quality assessment using RoB2 and ROBINS-I tools, the two independent reviewers reached a substantial agreement [59,60] During the selection process, we came across to some publications assessing the impact of the vaccine on RRP and HNC, but they failed to meet the defined inclusion criteria.
The limitations of our review mainly arise from the limitations of the included studies. Overall, the studies included males of a narrow range of age, mostly young males with somewhat limited sexual activity, thereby challenging the generalization of the results to other populations [17,31,32,37,42]. On the other hand, several studies analyzed small population samples from the outset, while others reported a smaller population size at the end of their studies, as dictated by discontinuation [34,38,40,41]. Most of the studies had short follow-up times, and no meaningful conclusions could be carried out for HPV-related premalignant and malignant lesions. We also noticed that for genital outcomes, most of the participants were heterosexual men, possibly leading to an underestimation of the vaccine efficacy itself, since these participants could benefit from herd immunity. Finally, the vaccine efficacy in the existing studies was expressed using absolute risk reductions, relative risk reductions, and a hazard ratio, and sometimes the incidence rate is the sole result. It became difficult and sometimes impossible to compare results and analyze the magnitude of the variations in efficacy among studies, which could have helped to assess their true significance. Additional limitations of our systematic review may also arise from the restriction of the outcomes to the clinical manifestations only. Our query could be made more inclusive by including synonyms of efficacy. However, reviewing all the references in the literature, we have not found any additional eligible study meeting the inclusion and exclusion criteria. A further limitation concerns the strict inclusion and exclusion criteria, but as stated at the beginning of our study, we aimed for high-quality evidence of the efficacy of HPV vaccination on male disease, not including immunogenicity studies.

Conclusions
HPV-related diseases in the male population represent an emerging problem, resulting in morbidity and mortality and associated with huge health costs due to recurrence and cancer risk. Finding a way to prevent them is a crucial public health strategy. Increasing vaccination coverage, regardless of sexual orientation or gender identity, and monitoring the impact of vaccination should become public health priorities.
This systematic review supports the recommendation of early vaccination of boys before the onset of sexual activity. There is clear evidence of the HPV vaccine's efficacy in men up to the age of 26 years old, with or without history of HPV-related disease. HPV vaccination protects against anal and genital warts and AIN1|2|3.
It is worth considering the impact of the HPV vaccine on HPV-related penile cancer and PIN in future studies. Moreover, in the future, it will be important to carry out RCTs that include more MSM and PLWH, and to evaluate the efficacy of vaccination on genital disease. An unanswered question in this systematic review involves the vaccine's efficacy against RRP and HNC; this is due to the lack of RCT studies addressing this topic.